Detection of Human Dystroglycan by Western Blot.
Western blot shows lysates of MCF‑7 human breast cancer cell line, SH‑SY5Y human neuroblastoma cell line, human muscle tissue, and human placenta tissue. PVDF membrane was probed with 1 µg/mL of Sheep Anti-Human Dystroglycan Antigen Affinity-purified Polyclonal Antibody (Catalog # AF6868) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). Specific bands were detected for alpha ‑Dystroglycan at approximately 100-160 kDa (as indicated) and beta ‑Dystroglycan at approximately 42-44 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
Dystroglycan in Human Skeletal Muscle.
Dystroglycan was detected in immersion fixed paraffin-embedded sections of human skeletal muscle using Sheep Anti-Human Dystroglycan Antigen Affinity-purified Polyclonal Antibody (Catalog # AF6868) at 3 µg/mL overnight at 4 °C. Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using the Anti-Sheep HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS019) and counterstained with hematoxylin (blue). Specific staining was localized to basement membrane. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.
Preparation and Storage
Sterile PBS to a final concentration of 0.2 mg/mL.
Reconstitution Buffer Available
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
Stability & Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
12 months from date of receipt, -20 to -70 °C as supplied.
1 month, 2 to 8 °C under sterile conditions after reconstitution.
6 months, -20 to -70 °C under sterile conditions after reconstitution.
Dystroglycan, also DAG-1 (Dystrophin-associated glycoprotein 1) is a 180-200 kDa heterodimeric adhesion molecule that links the cell cytoskeleton to the extracellular matrix. It is found on skeletal muscle, cardiac muscle, fibroblasts, smooth muscle and keratinocytes. DAG-1 binds multiple matrix molecules, including laminin-1 and -2, agrin, and perlecan. Intracellularly, the cytoplasmic tail of DAG-1 contributes to a large 400 kDa complex that interacts with the cytoskeleton. The human DAG-1 preprocursor is a type I transmembrane protein 895 amino acids (aa) in length. It contains a 27 aa signal sequence plus an 868 aa proform that undergoes autocatalysis to generate a 626 aa alpha -chain (aa 28-653), and a 242 aa beta -chain. Mature DAG-1 is a heterodimer composed of noncovalently linked alpha ‑ and beta ‑chains. The alpha -chain possesses one potential Ig-like domain (aa 64-162), a mucin-like region (aa 316-485), and a peptidase S72 domain (aa 500-733). It is O‑glycosylated and runs from 100-160 kDa in SDS-PAGE. The beta -chain is N-glycosylated and runs at 42-44 kDa in SDS-Page. It possesses a short 95 aa extracellular region (aa 654-749) plus a 120 aa cytoplasmic domain (aa 776-895). Membrane cleavage of the beta -chain causes dissociation of the heterodimer and generates a 30 kDa truncated form. Over aa 28-749, human DAG-1 shares 93% aa identity with mouse DAG-1.
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